This disclosure pertains to light-emitting nanoparticles and their uses in biological imaging and sensing applications.
Development of light-emitting polymer nano/micro particles is usually hampered by inherent drawbacks such as photobleaching, stability, and functionalization issues resulting from fluorescent/dye dopants. Formation of biocompatible, luminescent polymer particles often entails hazardous chemical cross-linking processes and/or doping with fluorophores susceptible to leaching and photobleaching.
Growing research interests over the usage of luminescent nanomaterials for diverse biomedical and materials applications have led to the development of various approaches to overcome underlying drawbacks. Numerous efforts are ongoing to address limitations in brightness, biocompatibility, bioconjugation, functionalization, solubility, and/or stability so as to bring the light-emitting material closer to practicality for each particular application. Doping and chemically-conjugating a light-emitter to a host matrix (e.g., polymer, silica/aluminosilica sol-gel matrix, or a carbon nanotube) represent two popular approaches that have met excellent success for enhancing the solubility and stability of luminescent systems. Nevertheless, issues related to guest leakage, complexities in nanoparticle formulations, and/or toxicity of cross-linkers necessitate further research for developing alternative materials and more facile techniques. Polyelectrolytic self-assembly techniques are advantageous in terms of simplicity for synthesizing a variety of nanostructures.
Fluorescent polyelectrolyte structures are considered advantageous compared to dye doped particles for minimizing diffusion or dissolution of dye. However, common fluorescent polyelectrolytic systems employed continue to suffer poor selectivity, photostability, and low quantum efficiency. Fluorescent systems based on pure organic moieties are liable to quenching when interfaced with polymer or host matrices compared to organometallic systems. Phosphorescent systems offer numerous advantages over fluorescent analogues, such as higher efficiencies in electroluminescence (devices based on phosphorescence can exhibit 4× higher efficiency compared to fluorescent ones possessing the same photoluminescence quantum yield), hypoxia sensing, amelioration of background interference (e.g., autofluorescence by endogenous biomolecules and host matrices) by time-based analysis, all of which can be major obstacles in different applications with fluorescence-based systems.
Well-known phosphorescent molecular systems based on Ru(II) or Ir(III) have been utilized in material design through doping techniques; however, the rational engineering of luminescent nanoparticles remains an elusive goal. There has been significant interest in utilizing benign linear polymers such as chitosan in the synthesis of advanced functional materials for biomedical applications because of its well-established biocompatibility and aqueous solubility. Water-soluble polymer systems are also advantageous for materials applications, such as solution processing of multi-layered polymer light-emitting diodes (PLEDs) and electronically-active conductive hydrogels. In fact, the use of both water- and organic-processed layers has proved to be an excellent method for depositing different electronic layers with minimal interference of the others. Whereas all-organic-media-processed devices can lead to interlayer mixing, degradation, and/or other parasitic effects—due to the layers having similar solubilites in the same organic or aqueous phase—alternating water-processed and organic-processed layers helps minimize these issues due to the inherent lower material solubility among successively-deposited layers. For instance, one could deposit a hole-injecting layer (HIL) from an aqueous phase, followed by a hole-transporting (HTL) and/or electron-blocking (EBL) layer from organic media, such as toluene or chlorobenzene, then a light-emitting layer (EML) of the type described in this work from aqueous solution, and finally an electron-transporting layer (ETL) from organic media—if needed (given the dual EML/ETL function of some compositions herein that may preclude the need for a separate ETL). The incompatible solubility between adjacent layer materials in organic vs. aqueous media or phases minimizes invasive interactions between the two deposited layers, hence warranting exciton and/or polaron confinement in the designated layer function(s).
Among existing techniques for synthesizing microparticles and nanoparticles from chitosan polysaccharides and other linear polymers, polyelectrolyte complexation or ionic gelation processes are highly preferred for their simplicity and because they do not rely on toxic chemical cross-linkers. Categorically, past efforts at incorporating luminescent moieties into chitosan nanomaterials have focused on doping or chemically-cross-linking fluorophores including organic dyes, such as fluorescein isothiocyanate (FITC), semiconductor quantum dots, or phosphorescent lanthanide chelates. In all these cases, however, chitosan was used as a capping agent or surfactant to overcome the toxicity or insolubility of luminophores and their incorporation required multiple steps. For example, studies detailing the synthesis of fluorescent chitosan nanoparticles using FITC-labeled chitosan employed a multistep microemulsion technique that included the non-luminescent cross-linkers 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) and tartaric acid.
Though chemical crosslinking will allow for the formation of micro/nanoparticles, during this process of chemical crosslinking the inherent properties of polymers are sacrificed due to the presence of additional chemical entities. Introducing additional cross-linkers or additives are also known to compromise the optical properties of fluorescent systems, resulting in sub-standard formulations. These drawbacks arise from chemical bonding between the host matrices and the fluorescent dyes that could be easily avoided by polyelectrolyte complexation, which completely relies on electrostatic interactions between oppositely-charged luminescent moieties and host matrix materials. The strength of ionic or physical cross-linking interactions can be easily tuned by variations in pH or ionic strength of the medium in order to favor weaker or stronger interactions that would then allow for size tunability of the polymer particles.